CAJ | 학술논문

目的通过改变部分翻译起始区序列观察对EPO模拟肽基因8串联体表达的影响.方法设计和人工合成了部分翻译起始区域（translationinitiation region,TIR）DNA序列，将该序列插入pBSEPOM8（含EPO模拟肽基因8串联体序列）的EcoRI和XbaI位点，经DNA测序确定正确重组质粒，再将EPO模拟肽基因8串联体（含改建的翻译起始区域）克隆到pBV220载体的EcoRI和BamHI位点，诱导表达.结果 DNA测序证明，人工合成的DNA序列已正确插入EPO模拟肽基因8串联体的翻译起始区域.该串联体基因插入pBV220载体后，经42℃诱导，在细菌裂解上清中出现一条相对分子质量为22×103(Mr)的新蛋白带，与EPO模拟肽基因8串联体的理论计算分子量相符合.经薄层扫描证明该蛋白带约占细菌蛋白总量的15%.结论通过改变部分翻译起始区域起动了EPO模拟肽基因8串联体的翻译，使原来不表达的EPO模拟肽基因8串联体获得了较高水平的表达.
목적 통과개변부분번역기시구서렬관찰대EPO모의태기인8천련체표체적영향.방법 설계화인공합성료부분번역기시구역（translationinitiation region,TIR）DNA서렬，장해서렬삽입pBSEPOM8（함EPO모의태기인8천련체서렬）적EcoRI화XbaI위점，경DNA측서학정정학중조질립，재장EPO모의태기인8천련체（함개건적번역기시구역）극륭도pBV220재체적EcoRI화BamHI위점，유도표체.결과 DNA측서증명，인공합성적DNA서렬이정학삽입EPO모의태기인8천련체적번역기시구역.해천련체기인삽입pBV220재체후，경42℃유도，재세균렬해상청중출현일조상대분자질량위22×103(Mr)적신단백대，여EPO모의태기인8천련체적이론계산분자량상부합.경박층소묘증명해단백대약점세균단백총량적15%.결론 통과개변부분번역기시구역기동료EPO모의태기인8천련체적번역，사원래불표체적EPO모의태기인8천련체획득료교고수평적표체.
AIM To observe effect of a synthesized DNA sequence oftranslation initiation region (TIR) on expression of 8 repeats of EOP mimetic peptide gene. METHODS A DNA sequence of translation initiation region was designed, synthesized and inserted into EcoRI and XbaI sites of pBSEPOM8 plasmid DNA was named for pBSEPOM8B. Then 8 repeats of EPO mimetic peptide gene which contained synthesized TIR sequence was cloned into EcoRI and BamHI sites of pBV220 expression vector. RESULTS DNA sequencing analysis showed that the sequence of synthesized TIR was correctly inserted into EcoRI and XbaI sites of pBVEPOM8B. After recumbent bacterium contained pBVEPOM8B was induced at 42℃ for 4 h, a new protein band was found on SDS-PAGE gel. The relative molecule mass of the new protein band was about 22 000 (Mr), consistent with 8 repeats of EPO mimetic peptide. The protein band amounted to 15% of total bacteria protein. CONCLUSION Synthesized DNA sequence of TIR initiates translation of 8 repeats of EPO mimetic peptide gene, and so helps improve the previously non-expression of the latter.